Chromatography is a powerful separation technique widely used in various scientific disciplines, including many pharmaceutical, chemical, and biotechnological analyses and preparative processes. In general, chromatography embraces a diverse collection of techniques for separating closely related components of complex mixtures by passing a mobile phase, which contains the sample to be separated, through an immiscible stationary phase. Mobile and stationary phases are selected such that sample constituents distribute themselves between the two phases to varying degrees. Each individual substance moves as a zone progressing at a fraction of the mobile phase rate. One constituent separates from others because this fraction varies according to the particular substance, depending on their partition coefficients between the two phases. Those constituents that strongly interact with, or adsorb on, the stationary phase advance at slower rates, if at all, than those with weaker molecular interactions do, thus effecting separation. The separated constituents are then accessible for subsequent downstream analysis.
Various characteristics provide a basis for separating materials. For example, ion chromatography generally involves the separation of sample constituents based upon their net charge. This sensitive technique is frequently used to separate organic or inorganic ions and even nonionic substances. It typically entails flowing the mobile phase through an ion exchanger having a net charge opposite from components to be separated from a sample mixture whether it is the material of interest or impurities and subsequently eluting the components from the exchanger. Other chromatographic techniques exploit differences, other than net charge or polarity, among sample components, such as distinguishing binding affinities for a selected stationary phase, and hydrophilic or hydrophobic characteristic variations, among other properties.
Many different chromatographic techniques are generally known and described in the literature, including, e.g., Matejtschuk (Ed.), Affinity Separations: A Practical Approach (1997) IRL Press, Oxford; Scouten, Affinity Chromatography: Bioselective Adsorption on Inert Matrices (1981) John Wiley & Sons, New York; Bickerstaff (Ed.) Immobilization of Enzymes and Cells: Methods in Biotechnology 1 (1997) Humana Press, Towana, N.J.; Hermanson et al., Immobilized Affinity Ligand Techniques (1992) Academic Press, San Diego; Hydrophobic Interaction Chromatography: Principles and Methods (1993) Pharmacia; Brown, Advances in Chromatography (1998) Marcel Dekker, Inc., New York; Fallon, Booth, and Bell (Eds.), Applications of HPLC in Biochemistry: Laboratory Techniques in Biochemistry and Molecular Biology (1987) Elsevier Science Publishers, Amsterdam; Lough and Wainer (Eds.), High Performance Liquid Chromatography: Fundamental Principles and Practice (1996) Blackie Academic and Professional, London; Mant and Hodges (Eds.), High Performance Liquid Chromatography of Peptides and Proteins: Separation, Analysis and Conformation (1991) CRC Press, Boca Raton; Katz (Ed.), High Performance Liquid Chromatography: Principles and Methods in Biotechnology (1996) John Wiley & Sons, Inc., Chichester, England; Weiss, Ion Chromatography, 2nd ed. (1995) VCH, New York; Ion-Exchange Chromatography: Principles and Methods (1991) Pharmacia; Smith, The Practice of Ion Chromatography (1990) Krieger Publishing Company, Melbourne, Fla.; and Bidlingmeyer, Practical HPLC Methodology and Applications (1992) John Wiley & Sons, Inc., New York.
In general, additional chromatographic techniques would be desirable. The present invention provides new methods and devices for performing chromatographic separations that have many significant advantages over current separation approaches. These and a variety of additional features will become apparent upon complete review of the following description.